Caitlin M Rodriguez1,2, Sang Y Chun3, Ryan E Mills4,5, Peter K Todd6,7. 1. Department of Neurology, University of Michigan, Ann Arbor, MI, USA. 2. Neuroscience Graduate Program, University of Michigan, Ann Arbor, MI, USA. 3. Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, MI, 48109, USA. 4. Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, MI, 48109, USA. remills@umich.edu. 5. Department of Human Genetics, University of Michigan, Ann Arbor, MI, 48109, USA. remills@umich.edu. 6. Department of Neurology, University of Michigan, Ann Arbor, MI, USA. petertod@med.umich.edu. 7. VA Ann Arbor Healthcare System, Ann Arbor, MI, USA. petertod@med.umich.edu.
Abstract
BACKGROUND: Upstream open reading frames (uORFs) initiate translation within mRNA 5' leaders, and have the potential to alter main coding sequence (CDS) translation on transcripts in which they reside. Ribosome profiling (RP) studies suggest that translating ribosomes are pervasive within 5' leaders across model systems. However, the significance of this observation remains unclear. To explore a role for uORF usage in a model of neuronal differentiation, we performed RP on undifferentiated and differentiated human neuroblastoma cells. RESULTS: Using a spectral coherence algorithm (SPECtre), we identify 4954 consistently translated uORFs across 31% of all neuroblastoma transcripts. These uORFs predominantly utilize non-AUG initiation codons and exhibit translational efficiencies (TE) comparable to annotated coding regions. On a population basis, the global impact of both AUG and non-AUG initiated uORFs on basal CDS translation were small, even when analysis is limited to conserved and consistently translated uORFs. However, uORFs did alter the translation of a subset of genes, including the Diamond-Blackfan Anemia associated ribosomal gene RPS24. With retinoic acid induced differentiation, we observed an overall positive correlation in translational shifts between uORF/CDS pairs. However, CDSs downstream of uORFs show smaller shifts in TE with differentiation relative to CDSs without a predicted uORF, suggesting that uORF translation buffers cell state dependent fluctuations in CDS translation. CONCLUSION: This work provides insights into the dynamic relationships and potential regulatory functions of uORF/CDS pairs in a model of neuronal differentiation.
BACKGROUND: Upstream open reading frames (uORFs) initiate translation within mRNA 5' leaders, and have the potential to alter main coding sequence (CDS) translation on transcripts in which they reside. Ribosome profiling (RP) studies suggest that translating ribosomes are pervasive within 5' leaders across model systems. However, the significance of this observation remains unclear. To explore a role for uORF usage in a model of neuronal differentiation, we performed RP on undifferentiated and differentiated humanneuroblastoma cells. RESULTS: Using a spectral coherence algorithm (SPECtre), we identify 4954 consistently translated uORFs across 31% of all neuroblastoma transcripts. These uORFs predominantly utilize non-AUG initiation codons and exhibit translational efficiencies (TE) comparable to annotated coding regions. On a population basis, the global impact of both AUG and non-AUG initiated uORFs on basal CDS translation were small, even when analysis is limited to conserved and consistently translated uORFs. However, uORFs did alter the translation of a subset of genes, including the Diamond-Blackfan Anemia associated ribosomal gene RPS24. With retinoic acid induced differentiation, we observed an overall positive correlation in translational shifts between uORF/CDS pairs. However, CDSs downstream of uORFs show smaller shifts in TE with differentiation relative to CDSs without a predicted uORF, suggesting that uORF translation buffers cell state dependent fluctuations in CDS translation. CONCLUSION: This work provides insights into the dynamic relationships and potential regulatory functions of uORF/CDS pairs in a model of neuronal differentiation.
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